Intracutaneous Injection Research Articles

Effect of Xinfeng Capsule on lung function in rats with adjuvant-induced arthritis and its mechanism

To investigate the effects of Xinfeng Capsule (XFC) on pulmonary function and related mechanism in adjuvant-induced arthritis (AA) rats. The rats were randomly divided into five groups: normal control (NC), model control (MC) groups, methotrexate (MTX), tripterygium glycosides tablet (TPT) and Xinfeng capsule (XFC) treatment groups. The adjuvant-induced arthritis model was established by intracutaneous injection of 0.1 mL Freund ' s complete adjuvant in the right paw of rats; the drugs were given 19 d after model establishment. The toe swelling degree (E), arthritis index (AI), pulmonary function, peripheral blood Treg levels, pathological changes of lung tissue and expression of Foxp3, TGF-β1, Smad3, Smad7 proteins in lung tissue were measured 30 d after drug administration. Compared to NC group, the levels of E, AI, alveolitis score, TGF-β1 and Smad3 were significantly increased (P <0.05 or P <0.01); maximum expiratory flow 25% of vital capacity (FEF25),50% maximal expiratory vital capacity flow (FEF50), maximum expiratory flow at 75% of vital capacity (FEF75), maximum mid-expiratory flow (MMF), force peak expiratory flow (PEF), CD4+ CD25+ Treg, Foxp3 and Smad7 were significantly decreased in MC group (P <0.05 or P < 0.01). Compared to MC group,the expression of E, AI, TGF-β1 and Smad3 were reduced, while FEF50, FEF75, MMF, PEF, Treg, Foxp3 and Smad7 were elevated in XFC group (P <0.05 or P <0.01). Compared to XFC group, the level of body mass,FEF25,FEF50, FEF75, MMF and Treg were lower in MTX and TPT groups (P <0.05 or P <0.01). There are inflamed joints and reduced pulmonary function in rats of adjuvant-induced arthritis. XFC can inhibit paw edema degrees, reduce arthritis response, and improve pulmonary function, which is associated with up-regulating expression of Treg and Foxp3, down-regulating the expression of TGF-β1 and adjusting TGF-β1/Smads signal pathway.

Assessment of surgical sutures Polymed® by intracutaneous irritation test in rabbits

The aim of the present study was to evaluate local irritant effects to rabbit skin following a single application of test samples of non-sterile polyamide non-absorbable surgical sutures POLYMED®. The polar and nonpolar extracts were prepared by using saline solution and olive oil, respectively, after sinking the materials tested (2.0 g) in 10 ml of the corresponding liquid. Incubation was carried out at the temperature of 37 °C for 72 h. The saline solution and pure olive oil, which had no contact with the materials tested, were used as negative control samples and were incubated under the same conditions as above. Assessments of the extracts from each material were conducted on 2 albino rabbits of the New Zealand breed. On the back of each animal, 5 intracutaneous injections of the extract tested and 5 injections of the control solution, each of 0.2 ml, were carried out. The degree of irritation was scored at 4, 24, 48, 72 hours after injection and no skin changes were found. The intracutaneous irritation index (III) was calculated and yielded 0.0. Hence it was concluded that under the experimental conditions the extracts of the material tested, i.e. non-sterile polyamide non-absorbable surgical sutures POLYMED®, were ‘non-irritant’ to the skin of rabbits when compared with the respective control groups. The experimental procedure was conducted according to ISO10993-10.

Local nociceptor sensitization with NGF: Mechanical or heat hyperalgesia á la carte?

In this issue, you will find a paper by C. Mills et al., (2013) entitled ‘Characterization of nerve growth factor induced mechanical and thermal hypersensitivity in the rat’. Nerve growth factor (NGF) has been found to sensitize pain processing already in the 1990s (Lewin et al.,1992; Woolf et al.,1994; Bennett et al.,1998; Koltzenburg et al.,1999; Kerr et al.,2001). Successful pain treatment using NGF antibodies (Lane et al.,2010; Katz et al.,2011; Schnitzer et al.,2011) has renewed the interest in this mechanism also inciting studies on human pain models applying NGF intramuscularly (Andersen et al.,2008; Svensson et al.,2008), in the muscle fascia (Deising et al.,2012) and in the skin of human volunteers (Rukwied et al.,2010; Weinkauf et al.,2012). Upon injection in the skin, NGF-induced mechanical and thermal hyperalgesia is found in animal models (Andreev et al.,1995). There are however, major differences in the time course of the hyperalgesia between tissues and between species. While the duration of hyperalgesia upon injection in the muscle is only a few days (Andersen et al.,2008; Svensson et al.,2008), intracutaneous injection in humans causes mechanical hyperalgesia lasting more than a month (Rukwied et al.,2010; Weinkauf et al.,2012). In rodents, heat and mechanical hyperalgesia have been reported to start hours after the injection and last hours or up to a couple of days. Mills and colleagues in this issue of European Journal of Pain report on a differential time course of heat and mechanical hyperalgesia in a rat model of NGF-induced hypersensitivity. Upon dermal injection of NGF, heat hyperalgesia was maximum after 3 h and had returned to baseline at 48 h. In contrast, mechanical hyperalgesia as measured with paw withdrawal threshold had developed 1 h after the injection but was significant for 1 week even for low NGF doses (0.3 μg) and for about 2 weeks at 3 μg. Interestingly, hypersensitivity to pin prick stimulation was significant only for the time point 1 h after the injection. The basis for the mismatch between pin prick hyperalgesia and withdrawal thresholds is unclear. However, one might suggest that the phasic pin prick tests are assessing behavioural pain threshold whereas the tonic nature of the paw pressure tests would allow more for suprathreshold testing. Along these lines, it is conceivable that the NGF-induced long-lasting mechanical hyperalgesia is based on sensitization of suprathreshold encoding as has similarly been found in the human NGF model (Rukwied et al.,2010), but also in the rat ultraviolet irradiation model (Bishop et al.,2010) in which high threshold mechanosensitive C-nociceptors were found to increase their firing upon suprathreshold mechanical stimulation, with their activation threshold being unchanged. The differential time course of mechanical and heat hypersensitivity resembles the delayed mechanical hyperalgesia observed in human volunteers and suggests different underlying mechanisms. While heat hyperalgesia might be based on sensitization and translocation of TRPV1, the prolonged mechanical sensitization might be linked to expression changes of axonal or sensory transduction proteins. The key implication of these results in respect to pharmacological testing is obviously the use of early and late hyperalgesia for separate testing of analgesic effects. The centrally acting analgesics morphine and gabapentin did not show a clear separation of their analgesic potency when comparing early and late phase or heat versus mechanical hyperalgesia. In contrast, duloxetin appeared to be more effective against the late mechanical hyperalgesia whereas the anti-inflammatory drugs Diclofenac and Celecoxib might be more effective in the early phase. In summary, the prolonged mechanical hyperalgesia as compared to the more transient heat hyperalgesia in the rat NGF model might allow for a better characterization of analgesic effects. Conflict of interest None declared.

Intracutaneous sterile water injection versus oral paracetamol for renal colic during pregnancy: a randomized controlled trial

The aim of this study was to assess the analgesic efficacy of intracutaneous sterile water injection compared with oral paracetamol in pregnant women with acute renal colic caused by urolithiasis. The study included 45 patients randomized into two different groups: Group A (n = 24) received paracetamol (1,000 mg, oral), while group B (n = 21) received sterile water injections. The severity of pain was assessed by a visual analogue scale (VAS) system at baseline and at 15, 30 min, and 1 h after administering the treatments. Subjects with inadequate pain relief at 1 h received rescue analgesia. The VAS values prior to the start of therapy and 15, 30 min, and 1 h after therapy were 85.42 ± 10.62, 69.17 ± 8.3, 45.42 ± 12.5, and 32.08 ± 14.44 for the paracetamol group, while for the sterile water injection group, the VAS values were 90.48 ± 11.17, 30.95 ± 16.7, 14.76 ± 11.23, and 10.48 ± 8.65, respectively. There was no statistically significant difference between the starting VAS values of the two groups, but the VAS values 15, 30 min, and 1 h after paracetamol administration were statistically less in the group that received sterile water injections. Rescue analgesics at 1 h were required by eight subjects (33 %) receiving paracetamol and one subject (5 %) receiving sterile water injection. Intracutaneous injection of sterile water is an efficacious treatment for renal colic caused by urolithiasis in pregnant women. It was significantly superior to oral administration of paracetamol.

Diagnosis of Inguinal Lymph Node Metastases Using Contrast Enhanced High Resolution MR Lymphangiography

Inguinal lymph nodes can be the first or the only clinical signs of tumor metastases. The aim of the study was to evaluate the role of contrast-enhanced high-resolution magnetic resonance (MR) lymphangiography in diagnosis of inguinal lymph node metastases. The study enrolled 26 patients with inguinal lymph node metastases. Contrast-enhanced lymphangiography was performed using a 3.0T MR unit after intracutaneous injection of gadobenate dimeglumine into the interdigital webs of the dorsal foot. Images of inguinal lymph nodes were acquired before and after contrast injection. All patients exhibited edema in the subcutaneous layer with significant dilatation of lymphatic collectors in the affected lower limbs on MR images. Before contrast injection, the outline and structure of the affected nodes were unclear on T2 weighted images. Structural changes became evident on postinjection T1-weighted images. Nodal involvement on contrast enhanced MR lymphangiograms was characterized as: 1) heterogeneous structure with partial or marginal enhancement of the node indicating partial occupation by tumor; 2) homogeneous structure of the node without contrast enhancement, indicating total occupation with metastasis, with increase or no change in size; and 3) heterogeneous structure with punctiform nodal enhancement indicating diffuse growth of tumor within the node. Further examinations confirmed the diagnoses of inguinal lymph node metastases of either regional or distal tumors. Contrast-enhanced high-resolution MR lymphangiography was a sensitive modality in the diagnosis of malignant peripheral lymphedema and the identification of inguinal lymph node metastasis in patients with various tumor origins.

An open-label prospective clinical study of the efficacy and safety of botulinus toxin type A for the correction of age-related neck skin changes

Objective. Assessment of the efficacy and safety of the correction of age-related neck skin changes with the use of botulinus toxin type A (Xeomin). Materials and methods. The study involved 25 women aged 39—65 enrolled according to the inclusion and exclusion criteria. The patients were administered a single subcutaneous injection of Xeomin 60 U in the neck. The efficacy and safety of this procedure was assessed based on the Global Aesthetic Improvement Scale (GAIS) on Day 14 as well as after 2 and 3 months. Results. High efficacy and safety of intracutaneous Xeomin injections at the submaximum dose of 60 u in the medial and lateral platysma bands have been confirmed clinically. The authors suggest an experimental gradining of the age-related neck making it possible to assess and statistically process clinical results of the new method.

Effects of Xinfeng capsule on pulmonary function based on treg-mediated notch pathway in a rat model of adjuvant arthritis

ObjectiveTo observe the impact of xinfeng xapsule (XFC) on pulmonary function in a rat model of adjuvant arthritis (AA) and to investigate the mechanism of action. MethodsForty rats were randomly divided into four groups of ten: normal control (NC); model control (MC); tripterygium glycosides tablet (TPT); and xinfeng capsule (XFC). Except for the NC group, AA was induced in all rats by intracutaneous injection of 0.1 mL Freund's complete adjuvant in the right paw on the 19th day. NC and MC groups were given (0.9%) physiological saline. The TPT and XFC groups were given TPT (10 mg/kg) and XFC (1.2 g/kg), respectively. Thirty days after administration, changes in paw edema (E), the arthritis index (AI), pulmonary function, levels of regulatory T-cells (Treg), ultrastructure of lung tissue, and expression of Notch receptors and ligands in lung tissue were observed. ResultsIn the MC group, E and the AI were increased and pulmonary function significantly decreased; the structure of alveolar type-II cells was damaged; ratios of Treg in peripheral blood were reduced; and expression of Notch receptors such as Notch3 and Notch4 and ligands such as Delta1 in lung tissue were significantly increased whereas expression of Notch1, Jagged1 and Jagged2 were significantly decreased. After intervention with XFC, E and the AI were decreased; pulmonary function was enhanced; the structure of alveolar type-II cells was improved; and expression of Treg, Notch1, Jagged1, Jagged2 was elevated, whereas that of Notch3, Notch4 and Delta1 was reduced. ConclusionXFC can not only inhibit E and the AI and improve joint symptoms, it can also improve pulmonary function and reduce inflammation in lung tissue. These actions could be carried out through increases in the expression of Treg, Notch receptors (Notch1) and ligands (Jagged1, Jagged2), and reductions in the expression of Notch3, Notch4 and Delta1. These phenomena would reduce the deposition of immune complexes and the inflammatory response in lung tissue, thereby improving joint symptoms and pulmonary function.

Anti-Tumor Immunity Elicited by Peptide Complex-Alpha-Fetoprotein and Glycoprotein 96

Purposes: To construct a recombinant vaccine alpha-fetoprotein (AFP)-glycoprotein (gp96) complex, and study its ability to induce specific CTL response and its protective effect against AFP-producing tumor. Material/Methods: A recombinant vaccine was constructed by conjugating mouse alpha-fetoprotein to glycoprotein 96. By way of intracutaneous injection, mice were primed and boosted with recombinant vaccine mAFP/gp96, whereas single mAFP or gp96 injection as controls. The ELISPOT and ELISA were used to measure the frequency of cells producing the cytokine IFN-gama in splenocytes and the level of anti-AFP antibody of serum from immunized mice respectively. In vivo tumor challenge was carried out to assess the immune effect of the recombinant vaccine. Results: By recombinant mAFP/gp96 vaccine immunization, the results of ELISPOT and ELISA showed that the number of splenic cells producing IFN-gama and the level of anti-AFP antibody of serum were significantly higher in mAFP/gp96 group than those in mAFP and gp96 groups (122.50±9.30 IFN-gama spots/106 cells vs 46.40±10.32 IFN-gama spots/106 cells, 12.14±7.33 IFN-gama spots/106 cells, P&lt;0.01; 164.52±11.22 µg/mL vs 56.32±8.23 µg/mL, 7.56±3.47 µg/mL, P&lt; 0.01). The tumor volume in mAFP/gp96 group was significantly smaller than that in mAFP and gp96 groups (32.46±6.35 mm3 vs 384.16±11.43 mm3, 832.54±12.72 mm3, P&lt; 0.01). Conclusions: The study further confirmed the function of glycoprotein 96s immune adjuvant. Sequential immunization with recombinant mAFP/gp96 vaccine could generate effective antitumor immunity on AFP-producing tumor. The recombined mAFP/gp96 vaccine may be suitable for serving as an immunotherapy for hepatocellular carcinoma.

Methodology for preclinical assay of pseudoallergy of injectable drugs (Ⅱ)—— rat model for assay of cutaneous pseudoallergy induced by injections

To develop animal models and methodologies for assay of pseudoallergy induced by injectable drugs. Rats cutaneous anaphylactoid reaction model was developed by intravenous injection of 0. 6% Evans blue(EB) followed by intracutaneous injection of test substance solutions 50 microL. Diameters of subcutaneous blue spots and EB exudation were assayed. Rat anaphylactoid reaction was characterized as vascular hyperpermeability which was measured by diameters of blue spots inside the skin and the EB exudation of the blue spots. Compound 48/80 caused severe bluing and EB exudation in the skin by inducing obvious vascular hyperpermeability which indicated that it can induce rat skin pseudoallergy. Normal saline or 5% glucose injection showed no obvious reactions. The rat pseudoallergy model was validated by intracutaneous injections of western drug injections and Chinese medicine. Rats could be developed into skin pseudoallergy model for preclinical safety evaluation of injectable drugs. The pseudoallergy reaction in this model is of high clinic consistency, sensitivity, reproducibility, and maneuverability. The model is suitable for the evaluation for pseudoallergy induced by injectable products prepared from Chinese materia medica This model can also be used for safety assay and quality control in manufacturing process, spot checking of marketed products, screening of allergen as well as studying of pseudoallergy mechanism.

Establishing aluminium contact allergy

Traditionally, contact allergy to aluminium has been established by patch testing with aluminium chloride hexahydrate in petrolatum at 2.0% and an empty Finn Chamber®. The aim of this study was to investigate different aluminium test preparations regarding an optimal compound and an optimal test concentration. Six different aluminium compounds and an empty Finn Chamber® were used to patch test 21 patients with aluminium contact allergy. Aluminium chloride hexahydrate in saline was used for intracutaneous injection of 19 patients. One test preparation, aluminium lactate 2.4%, was found to show significantly more positive patch test reactions than aluminium chloride hexahydrate 2.0% (p = 0.03). Aluminium chloride hexahydrate at 10.0% pet. gave the highest number of positive reactions to aluminium [14/21 (67%)]. No positive reactions were noted to an empty Finn Chamber®, and 3 of 19 (16%) patients reacted positively to the intradermal test. The results of this study indicate that patch testing with aluminium chloride hexahydrate 2.0%, with an empty Finn Chamber® and the intradermal test with the salt and doses used are insufficient methods to detect contact allergy to aluminium. Aluminium chloride hexahydrate at 10.0% gave the highest number of positive reactions to aluminium.

Fasudil, a Rho-kinase inhibitor, attenuates lipopolysaccharide-induced vascular hyperpermeability and colonic muscle relaxation in guinea pigs

BackgroundRho-associated coiled coil-forming protein kinase (Rho-kinase), a downstream target effector of the small GTP-binding protein Rho, plays a key role in cell adhesion, motility, and contraction. The goal of the present study was to determine the role of the Rho/Rho-kinase signal pathway in the pathogenesis of lipopolysaccharide (LPS)-induced vascular hyperpermeability using the Rho-kinase inhibitor fasudil. MethodsTo evaluate plasma leakage, fasudil (3 or 10 mg/kg) or saline was intravenously administered 30 min before LPS injection. LPS (100, 300, and 1,000 μg/0.1 mL/site) and saline (0.1 mL/site) were administered intracutaneously in the dorsum of guinea pigs. Vascular permeability was measured on the dorsal skin by the local accumulation of Evans Blue dye after intracutaneous injection of LPS (100–1000 μg/site) from Escherichia coli. For the measurement of colonic muscle tension, fasudil (3 mg/kg) or saline was intravenously administered 30 min before LPS injection. LPS (1 mg/kg) was administered intravenously. ResultsDye leakage in the skin increased significantly 2 h after the injection of LPS. This LPS-induced dye leakage was significantly suppressed by fasudil (3 and 10 mg/kg). LPS caused a transient decrease in colonic muscle tension, which peaked 2.5 h after the injection. This decrease in muscle tension was significantly suppressed by pretreatment with fasudil (3 mg/kg). ConclusionsThe Rho/Rho-kinase pathway might play an important role in the pathogenesis of LPS-induced endotoxemia, and fasudil could attenuate LPS-induced microvascular permeability, leading to inhibition of endotoxemia.

Blood glutamate scavengers prolong the survival of rats and mice with brain-implanted gliomas.

L-Glutamate (Glu) plays a crucial role in the growth of malignant gliomas. We have established the feasibility of accelerating a naturally occurring brain to-blood Glu efflux by decreasing blood Glu levels with intravenous oxaloacetate, the respective Glu co-substrate of the blood resident enzyme humane glutamate–oxaloacetate transaminase(hGOT). We wished to demonstrate that blood Glu scavenging provides neuroprotection in the case of glioma.We now describe the neuroprotective effects of blood Glu scavenging in a fatal condition such as brain-implanted C6 glioma in rats and brain-implanted human U87 MG glioma in nude mice. Rat (C-6) or human (U87) glioma cells were grafted stereotactically in the brain of rats or mice. After development of tumors, the animals were drinking oxaloacetate with or without injections of hGOT. In addition, mice were treated with combination treatment, which included drinking oxaloacetate with intracutaneous injections of hGOT and intraperitoneal injection of Temozolomide. Animals drinking oxaloacetate with or without injections of hGOT displayed a smaller tumor volume, reduced invasiveness and prolonged survival than control animals drinking saline. These effects were significantly enhanced by Temozolomide in mice, which increased survival by 237%. This is the first demonstration of blood Glu scavenging in brain cancer, and because of its safety, is likely to be of clinical significance for the future treatment of human gliomas. As we demonstrated, the blood glutamate scavenging treatment in combination with TMZ could be a good candidate or as an alternative treatment to the patients that do not respond to TMZ.

Blood glutamate scavengers prolong the survival of rats and mice with brain-implanted gliomas.

SummaryL-Glutamate (Glu) plays a crucial role in the growth of malignant gliomas. We have established the feasibility of accelerating a naturally occurring brain to-blood Glu efflux by decreasing blood Glu levels with intravenous oxaloacetate, the respective Glu co-substrate of the blood resident enzyme humane glutamate–oxaloacetate transaminase (hGOT). We wished to demonstrate that blood Glu scavenging provides neuroprotection in the case of glioma. We now describe the neuroprotective effects of blood Glu scavenging in a fatal condition such as brain-implanted C6 glioma in rats and brain-implanted human U87 MG glioma in nude mice. Rat (C-6) or human (U87) glioma cells were grafted stereotactically in the brain of rats or mice. After development of tumors, the animals were drinking oxaloacetate with or without injections of hGOT. In addition, mice were treated with combination treatment, which included drinking oxaloacetate with intracutaneous injections of hGOT and intraperitoneal injection of Temozolomide. Animals drinking oxaloacetate with or without injections of hGOT displayed a smaller tumor volume, reduced invasiveness and prolonged survival than control animals drinking saline. These effects were significantly enhanced by Temozolomide in mice, which increased survival by 237%. This is the first demonstration of blood Glu scavenging in brain cancer, and because of its safety, is likely to be of clinical significance for the future treatment of human gliomas. As we demonstrated, the blood glutamate scavenging treatment in combination with TMZ could be a good candidate or as an alternative treatment to the patients that do not respond to TMZ.

Intracutaneous or subcutaneous sterile water injection compared with blinded controls for pain management in labour

Intracutaneous or subcutaneous injection of sterile water is rapidly gaining popularity as a method of pain relief in labour and it is therefore essential that it is properly evaluated. Adequate analgesia in labour is important to women worldwide. Sterile water injection is inexpensive, requires basic equipment, and appears to have few side effects. It is purported to work for labour pain. To determine the efficacy of sterile water injections for relief of pain (both typical contraction pain and intractable back pain) during labour compared to placebo (isotonic saline injections) or non-pharmacological interventions, and to identify any relevant effects on mode and timing of delivery, or safety of both mother and baby. We searched the Cochrane Pregnancy and Childbirth Group's Trials Register (30 May 2011), MEDLINE, and EMBASE (January 2010 to 30 May 2011), together with reference lists in retrieved studies and review articles. We included randomised, double blind, controlled studies using intracutaneous or subcutaneous sterile water injections for pain relief during labour. There were no restrictions on birth place, parity, risk, age, weight, gestation, or stage of labour. Potential comparators were placebo (saline) and non-pharmacological interventions (e.g. hypnosis or biofeedback). Two review authors independently assessed eligibility and quality of trials, and extracted data. We resolved any disagreements or uncertainties by discussion with a third review author. Primary outcome measures were at least 50% pain relief, or at least 30%, pain relief, patient global impression of change of at least 'good', mode of delivery, perinatal morbidity and mortality, maternal complications and adverse events. Secondary outcomes were women with any pain relief, use of rescue analgesia, and treatment group average pain relief. We made explicit judgements about potential biases in the studies. We included seven studies, with 766 participants: four used intracutaneous injections, two subcutaneous, and one both. All reported on low back pain in labour only. Methodological quality was good, but four studies were at high risk of bias due to small size of treatment groups, incomplete outcome data, and performance bias.All studies reported treatment group mean or median scores, finding greater reduction in pain for sterile water. However, failure to demonstrate a normal distribution for pain intensity or relief, and use of different scales, meant meta-analysis was inappropriate. No study reported primary dichotomous efficacy outcomes. One reported the number self-scoring 4/10 cm or more reduction in pain; significantly more had this outcome with sterile water (50% to 60%) than with placebo (20% to 25%).There was no significant difference between sterile water and saline for rates of caesarean section (risk ratio (RR) 0.58, 95% confidence interval (CI) 0.33 to 1.02), instrumental delivery (RR 1.31, 95% CI 0.79 to 2.18), rescue analgesia (RR 0.86, 95% CI 0.44 to 1.69), timing of delivery, or Apgar scores. Two studies reported that more women treated with sterile water would request the same analgesia in future.No study reported on women's satisfaction with pain relief, women's sense of control in labour, women's satisfaction with the childbirth experience, mother/baby interaction, rates of breastfeeding, maternal morbidity, infant long-term outcomes, or cost. No adverse events were reported other than transient pain with injection, which was worse with sterile water. The outcomes reported severely limit conclusions for clinical practice. We found little robust evidence that sterile water is effective for low back or any other labour pain. Neither did we find any difference in delivery or other maternal or fetal outcomes. Further large, methodologically rigorous studies are required to determine the efficacy of sterile water to relieve pain in labour.

The Use of an Artificial Skin Model to Study Transdermal Absorption of Drugs in Inflamed Skin

Studies on drug disposition in inflamed skin are important for safe and effective application of topical drugs. Here, the absorption of flurbiprofen (FP) through inflamed skin was examined in vivo and in a skin-mimicking artificial model system. The model skin system consisted of a silicone membrane acting as a model stratum corneum, laminated dialysis membranes acting as a model of viable skin, and 2 microdialysis probes-one used for determination of FP concentration and one acting as a model vessel. This model system could be used for quantitative evaluation of complicated permeation processes. In the in vivo experiments, FP absorption was suppressed in rats with inflamed skin induced by an intracutaneous injection of a mixed solution of λ-carrageenan, zymosan, and casein. Bovine serum albumin solution was placed between the dialysis membranes in the model skin system to mimic protein leaching in skin; the results suggested that the delayed absorption of FP in inflamed skin was due to binding to serum proteins leaching in the tissue. Such a combination of in vivo experiments and a model skin system is useful for understanding complex phenomena in inflamed and damaged skin and reduces experimental animal use.

Specific Genetic Immunotherapy Induced by Recombinant Vaccine Alpha-Fetoprotein-Heat Shock Protein 70 Complex

Abstract Purposes: To construct a recombinant vaccine alpha-fetoprotein (AFP)-heat shock protein (HSP70) complex, and study its ability to induce specific CTL response and its protective effect against AFP-producing tumor. Material/Methods: A recombinant vaccine was constructed by conjugating mouse alpha-fetoprotein to heat shock protein 70. By way of intracutaneous injection, mice were primed and boosted with recombinant vaccine mAFP/HSP70, whereas single mAFP or HSP70 injection as controls. The ELISPOT and ELISA were used to measure the frequency of cells producing the cytokine IFN-γ in splenocytes and the level of anti-AFP antibody of serum from immunized mice respectively. In vivo tumor challenge were carried out to assess the immune effect of the recombinant vaccine. Results: By recombinant mAFP/HSP70 vaccine immunization, the results of ELISPOT and ELISA showed that the number of splenic cells producing IFN-γ and the level of anti-AFP antibody of serum were significantly higher in mAFP/HSP70 group than those in mAFP and HSP70 groups (108.50±11.70 IFN-γ spots/106 cells vs 41.60±10.40 IFN-γ spots/106 cells, 7.32±3.14 IFN-γ spots/106 cells, P

Effects of Chinese herbal medicine Xinfeng Capsule on expressions of platelet-activating factor and interleukins 6 and 17 in peripheral blood of rats with adjuvant arthritis

To observe the levels of platelet-activating factor (PAF), interleukin-6 (IL-6) and IL-17 in peripheral blood of rats with adjuvant arthritis (AA), and the effects of Xinfeng Capsule (XFC), a compound traditional Chinese herbal medicine. A total of 40 male Sprague-Dawley rats were randomized into normal control group, model group, methotrexate (MTX) group, Tripterygium Wilfordii polycoride Tablet (TPT) group, and XFC group, respectively. AA was induced in rats by intracutaneous injection of 0.1 mL Freund's complete adjuvant in the right hindlimb. Contents of PAF, IL-6 and IL-17 in peripheral blood were measured by enzyme-linked immunosorbent assay. Body weight of rats, paw swelling and arthritis index (AI) were also observed. Compared with the normal control group, the levels of PAF, IL-6 and IL-17 in peripheral blood, paw swelling and AI were significantly increased in the model group (P<0.01). Compared with the model group, levels of PAF, IL-6 and IL-17, paw swelling and AI of the three drug-treated groups were significantly decreased (P<0.01). The body weight of the XFC group was significantly higher than those of the MTX- and TPT-treated groups. Compared with the MTX- and TPT-treated groups, level of PAF in the XFC group was significantly increased (P<0.05). The level of PAF in peripheral blood of the AA rats was positively correlated with IL-6 and IL-17 levels in peripheral blood, paw swelling and AI (P<0.01). The expression of PAF increases in peripheral blood of rats with AA, and it correlates with IL-6, IL-17, paw swelling and AI. XFC can decrease the expression of PAF, restrain infection induced by the activation of platelets, decrease the levels of IL-6 and IL-17, and hence decrease the paw swelling of rats with AA.

Sentinel lymph node biopsy with a gamma probe in patients with high-risk cutaneous squamous cell carcinoma

Lymphoscintigraphy (LS) and sentinel lymph node biopsy have become the standard of care for melanoma and breast cancer. However, the data on patients with cutaneous squamous cell carcinoma (SCC) are limited. We aimed to evaluate and identify the role of LS and sentinel lymph node biopsy in patients with high-risk cutaneous SCC. Nineteen patients (13 men, six women; 47-87 years of age, mean age 67.5 ± 12.3) with SCC were included in the study. LS was performed on all patients after intracutaneous injection of Tc-99m nanocolloid. Primary lesions and sentinel lymph nodes (SLNs) were excised with the help of a gamma probe. A total of 26 SLNs and 32 secondary lymph nodes were imaged on LS and were marked. During surgery, 29 SLNs, 21 secondary lymph nodes and three nonactive lymph nodes were excised. In total, 53 lymph nodes were removed surgically. A histopathological study revealed that all lymph nodes were negative for metastasis. Patients were followed up for an average of 41.1 ± 22.2 months (7-80 months). Until the time of data collection, 14 patients were alive and had no regional lymph node or distant metastasis. Local recurrence was seen in only one patient. He was reoperated upon 38 months ago. The feasibility of determining SLNs using LS and an intraoperative gamma probe in patients with cutaneous SCC was shown. Unnecessary elective lymph node dissection and possible complications could be avoided in 19 patients.

Anti-Tumor Immunotherapy Elicited by Cross-Link Peptide Vaccine Alpha-Fetoprotein and Heat Shock Protein 70 Complex

Purposes: To construct a recombinant vaccine alpha-fetoprotein (AFP)-heat shock protein (HSP70) complex, and study its ability to induce specific CTL response and its protective effect against AFP-producing tumor. Material/Methods: A recombinant vaccine was constructed by conjugating mouse alpha-fetoprotein to heat shock protein 70. By way of intracutaneous injection, mice were primed and boosted with recombinant vaccine mAFP/HSP70, whereas single mAFP or HSP70 injection as controls. The ELISPOT and ELISA were used to measure the frequency of cells producing the cytokine IFN-gama in splenocytes and the level of anti-AFP antibody of serum from immunized mice respectively. In vivo tumor challenges were carried out to assess the immune effect of the recombinant vaccine. Results: By recombinant mAFP/HSP70 vaccine immunization, the results of ELISPOT and ELISA showed that the number of splenic cells producing IFN-gama and the level of anti-AFP antibody of serum were significantly higher in mAFP/HSP70 group than those in mAFP and HSP70 groups (108.50±11.70 IFN-gama spots/106 cells vs 41.60±10.40 IFN-gama spots/106 cells, 7.32±3.14 IFN-gama spots/106 cells, P&lt;0.01; 156.32±10.42 µg/mL vs 66.52±7.35 µg/mL, 5.73±2.89 µg/mL, P&lt;0.01). The tumor volume in mAFP/HSP70 group was significantly smaller than that in mAFP and HSP70 groups (42.44±7.14 mm3 vs 392.23±12.46 mm3, 838.63±13.84 mm3, P&lt;0.01). Conclusions: The study further confirmed the function of heat shock protein 70’s immune adjuvant. Sequential immunization with recombinant mAFP/HSP70 vaccine could generate effective antitumor immunity on AFP-producing tumor. The recombined mAFP/HSP70 vaccine may be suitable for serving as an immunotherapy for hepatocellular carcinoma.

Anhidrosis associated with hypothalamic lesions related to anti-aquaporin 4 autoantibody

Serum anti-aquaporin 4 (AQP4) antibody is detected with high specificity in patients with neuromyelitis optica (NMO), characterised by recurrent optic neuritis and longitudinally extensive transverse myelitis [1, 2]. Three NMO patients with anti-AQP4 antibody were reported to have hypothalamic lesions, showing reduced hypocretin-1 levels in cerebrospinal fluid (CSF) and symptomatic narcolepsy [3]. Although the hypothalamus and hypocretin-1 regulate autonomic functions as well as sleep, autonomic functions in NMO patients with hypothalamic lesions have not been described well. Here we report the case of a patient with anhidrosis and narcolepsy as initial symptoms of disorders related to anti-AQP4 antibody. A 41-year-old woman was experiencing excessive daytime sleepiness. Cataplexy was not evident. One month after, she became aware of anhidrosis on the left side of her body including the face. The following month, she was admitted to our hospital because of hypersomnia and anhidrosis that extended throughout her entire body. Physical and neurological examination revealed hypotension (93/62 mmHg) and dry skin. Brain magnetic resonance imaging (MRI) showed a high signal intensity area around the third ventricle on fluid-attenuated inversionrecovery image (Fig. 1a). Multiple sleep latency test showed shortening of the mean sleep latency (4.8 min; normal: [8 min) and sleep-onset REM periods. Lumbar puncture showed only a decrease of CSF hypocretin-1 levels (177 pg/ml, normal:[200 pg/ml). Serum anti-AQP4 antibody was positive. Although sympathetic skin responses (SSR) were absent in all limbs (Fig. 1c), intracutaneous acetylcholine injection caused normal sweating reaction. The patient was diagnosed with NMO spectrum disorders having narcolepsy and anhidrosis, secondary to hypothalamic lesions associated with anti-AQP4 antibody, and treated with intravenous methylprednisolone (1 g/day) for 3 days, followed by oral prednisolone (1 mg/kg/day). After treatment, her excessive sleepiness and anhidrosis disappeared in a few days. MRI showed improvement of the abnormal intensity area (Fig. 1b), and CSF hypocretin-1 levels had recovered to 213 pg/ml. Moreover, repeated tests of SSR showed positive responses (Fig. 1d). Anhidrosis of the entire body was detected and confirmed by SSR in our patient associated with anti-AQP4 antibody as well as symptomatic narcolepsy. After treatment, the reduced CSF hypocretin-1 levels returned to normal in parallel with improvement of excessive daytime sleepiness, similar to previously reported NMO cases [3]. The hypothalamus is the highest level of integration of autonomic function; tumors, trauma, inflammation, or vascular disorders in this structure are known to cause autonomic dysfunctions such as hypothermia or abnormal T. Sekiguchi S. Ishibashi T. Kubodera J. Fukabori T. Yokota (&) H. Mizusawa Department of Neurology and Neurological Science, Graduate School, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8519, Japan e-mail: tak-yokota.nuro@tmd.ac.jp